Synaptic transmission is mediated by release of fast-acting transmitters at synapses via synaptic vesicle (SV) exocytosis. Synaptic modulation is, in contrast, mediated by release from dense-core vesicles (DCVs) of modulatory transmitters that act pre- and postsynaptically to modify synaptic transmission. The understanding of the molecular mechanisms that underlie rapid Ca2+-dependent SV exocytosis has increased over the past decade and the same molecular constituents are required for DCV exocytosis. However, differences in the physiological regulation of SV and DCV exocytosis suggest that there are also molecular mechanisms distinct to each. We discovered that CAPS (ca2+-dependent activator protein for secretion) resides on DCVs but not SVs and is required for DCV but not SV exocytosis. In our continuation studies, we will determine the molecular mechanism through which CAPS acts to facilitate Ca2-dependent DCV fusion with the plasma membrane. This will be accomplished by identifying domains on CAPS that mediate its interactions with plasma and DCV membranes (Aim 1). These studies will define the basis of the DCV-selectivity of CAPS function. To further elucidate the mechanism of CAPS action, the functional importance of its interactions with proteins such as syntaxin, rabphilin and Muncl8 will be determined (Aim 2). These studies will provide a molecular explanation of how CAPS regulates the fusion machinery to facilitate DCV exocytosis. To further define domains of CAPS required for function, we will characterize loss-of-function CAPS mutants as well as determine CAPS structure by X-ray crystallographic studies (Aim 3). To relate the molecular interactions of CAPS to its role in DCV exocytosis, we will study fusion pore dynamics in cells with modified CAPS function (Aim 4). Completion of these studies will provide insight on the regulation of the fusion machinery and on molecular differences between DCV and SV exocytosis. The results may find application in the therapy of nervous system and endocrine disorders that involve hypo- or hypersecretion of monoamine transmitters or peptide hormones.